Comparison of methods for the determination of β-aspartylglycine in fecal supernatants of leukemic patients treated with antimicrobial agents

β-Aspartylglycine is an indicator of the absence of bacterial enzymatic activity in the intestinal tract. This study describes and compares four methods — (1) dansylation with thin-layer chromatography, (2) ion-exchange chromatography, (3) thin-layer electrophoresis, (4) high-voltage paper electrophoresis — to determine the concentration of β-aspartylglycine in fecal supernatants of leukemic patients treated with antimicrobial agents.     

A comparative study of ectonucleotidase and P2 receptor mRNA profiles in C6 cell line cultures and C6 <Emphasis Type="Italic">ex vivo</Emphasis> glioma model

Glioblastoma multiforme is the most common type of primary brain tumour and has the worst clinical outcome. Nucleotides represent an important class of extracellular molecules involved in cell proliferation, differentiation and apoptosis. Alterations in purinergic signalling have been implicated in pathological processes, such as cancer, and glioma cell lines are widely employed as a model to study the biology of brain tumours. Increasing evidence, however, suggests that glioma cell lines may not present all the phenotypic and genetic characteristics of the primary tumours. We have compared the biological characteristics of C6 rat glioma cells in culture and the same cells after their implantation in the rat brain and growth in culture (denominated as the C6 ex vivo culture model). Parameters evaluated included cell morphology, differentiation, angiogenic markers, purinergic receptors and ecto-nucleotidase mRNA profile/enzymatic activity. Analysis of the C6 glioma cell line and C6 ex vivo glioma cultures revealed distinct cell morphologies, although cell differentiation and angiogenic marker expressions were similar. Both glioma models co-expressed multiple P2X and P2Y receptor subtypes with some differences. In addition, the C6 glioma cell line and C6 ex vivo glioma cultures exhibited similar extracellular ATP metabolism and cell proliferation behaviour when exposed to cytotoxic ATP concentrations. Thus, the disruption of purinergic signalling is a feature shown not only by glioma cell lineages, but also by primary glioma cultures. Our results therefore suggest the participation of the purinergic system in glioma malignancy. This study was supported by grants from the Brazilian agencies CNPq, FAPERGS and Fundo de Incentivo à Pesquisa e Eventos (HCPA). E. Braganhol and D. Huppes were recipients of Brazilian CNPq fellowships; A. Bernardi was the recipient of a CAPES fellowship.     

Toward an Information Tool for Integrated Product Policy: Requirements for Data and Computation

Integrated product policy, according to the European Union, requires reliable data on the impact of consumer products along their life cycles. We argue that this necessarily requires the development of an information tool for hybrid analysis, combining aspects of life-cycle assessment and input-output analysis. A number of requirements in the development of such a hybrid information tool are identified, mainly concerning data and computational structure. For the former, some important points of attention are discussed, whereas for the latter, operational formulas are developed.     

Extensive Set of 16S rRNA-Based Probes for Detection of Bacteria in Human Feces

For the detection of six groups of anaerobic bacteria in human feces, we designed seven new 16S rRNA-based oligonucleotide probes. This set of probes extends the current set of probes and gives more data on the composition of the human gut flora. Probes were designed for Phascolarctobacterium and relatives (Phasco741), Veillonella (Veil223), Eubacterium hallii and relatives (Ehal1469), Lachnospira and relatives (Lach571), and Eubacterium cylindroides and relatives (Ecyl387), and two probes were designed for Ruminococcus and relatives (Rbro730 and Rfla729). The hybridization conditions for the new probes were optimized for fluorescent in situ hybridization, and the probes were validated against a set of reference organisms. The probes were applied to fecal samples of 11 volunteers to enumerate their target bacterial groups. The Phasco741 and Veil223 probes both detected average numbers below 1% of the total number of bacteria as determined with the bacterial kingdom-specific Bact338 probe. The Ecyl387 probe detected about 1.4%, the Lach571 and Ehal1469 probes detected 3.8 and 3.6%, respectively, and a combination of the Rbro730 and Rfla729 probes detected 10.3%. A set of 15 probes consisting of probes previously described and those presented here were evaluated in hybridization with the fecal samples of the same volunteers. Together, the group-specific probes detected 90% of the total bacterial cells.     

Environmental life cycle assessment of linoleum

Linoleum is a floor covering consisting mainly of linseed oil, other vegetable oils, wood flour and limestone on a carrier of jute. Forbo-Krommenie B.V. commissioned the Centre of Environmental Science (CML) to carry out an Environmental Life Cycle Assessment for linoleum floors. The goal of this study was to assess the environmental performance of linoleum floors, indicating possible options for improvement, and assessing the sensitivity of the results to methodological choices. The functional unit was defined as: 2000 m² linoleum produced in 1998, used in an office or public building over a period of 20 years. The method followed in this study is based on a nearly final draft version of the LCA guide published by CML in corporation with many others, which is an update of the guide on LCA of 1992. From the contribution analysis, the main contributing processes became clear. In addition, the sensitivity analysis by scenarios showed that the type of maintenance during use and the pigments used can have a large influence on the results. Major data gaps of the study were capital goods and unknown chemicals. Sensitivity analysis also showed that these gaps can lead to an underestimation. Based on this study, some options to improve the environmental performance of linoleum were formulated and advice for further LCA studies on linoleum was given.     

Missing inventory estimation tool using extended input-output analysis

Intention, Goal, Scope, Background  Input-Output Analysis (IOA) has recently been introduced to Life Cycle Assessment (LCA). In applying IOA to LCA studies, however, it is important to note that there are both advantages and disadvantages. Objectives  This paper aims to provide a better understanding of the advantages and disadvantages of adopting IOA in LCA, and introduces the methodology and principles of the Missing Inventory Estimation Tool (MIET) as one of the approaches to combine the strengths of process-specific LCA and IOA. Additionairy, we try to identify a number of possible errors in the use of IOA for LCA purposes, due to confusion between industry output and commodity, consumer’s price and producer’s price. Method  MIET utilises the 1996 US input-output table and various environmental statistics. It is based on an explicit distinction between commodity and industry output. Results and Discussion  MIET is a self-contained, publicly available database which can be applied directly in LCA studies to estimate missing processes. Conclusion  By adopting MILT results in existing, process-based, life-cycle inventory (LCI), LCA practitioners can fully utilise the process-specific information while expanding the system boundary. Recommendations and Outlook  MIET will be continuously updated to reflect both methodological developments and newly available data sources. For supporting information sec http:// wwwJeidenuniv.nl/cml/ssp/softwarc/miet.     

Enzyme optimization: moving from blind evolution to statistical exploration of sequence-function space

Directed evolution is a powerful tool for the creation of commercially useful enzymes, particularly those approaches that are based on in vitro recombination methods, such as DNA shuffling. Although these types of search algorithms are extraordinarily efficient compared with purely random methods, they do not explicitly represent or interrogate the genotype-phenotype relationship and are essentially blind in nature. Recently, however, researchers have begun to apply multivariate statistical techniques to model protein sequence-function relationships and guide the evolutionary process by rapidly identifying beneficial diversity for recombination. In conjunction with state-of-the-art library generation methods, the statistical approach to sequence optimization is now being used routinely to create enzymes efficiently for industrial applications.     

Environmental Impacts of Consumption in the European Union:High-Resolution Input-Output Tables with Detailed Environmental Extensions

For developing product policy, insight into the environmental effects of products is required. But available life-cycle assessment studies (LCAs) are hardly comparable between different products and do not cover total consumption. Input-output analysis with environmental extensions (EEIOA) of full consumption is not available for the European Union. Available country studies have a low sector resolution and a limited number of environmental extensions. This study fills the gap between detailed LCA and low-resolution EEIOA, specifying the environmental effects of household consumption in the European Union, discerning nearly 500 sectors, while specifying a large number of environmental extensions. Added to the production sectors are a number of consumption activities with direct emissions, such as automobile driving, cooking and heating, and a number of postconsumer waste management sectors. The data for Europe have been constructed by using the sparse available and coarse economic and environmental data on European countries and adding technological detail mainly based on data from the United States.
A small number of products score high on environmental impact per Euro and also have a substantial share of overall consumer expenditure. Several meat and dairy products, household heating, and car driving thus have a large share of the total environmental impact. Due to their sales volume, however, products with a medium or low environmental score per Euro may also have a substantial impact. This is the case with bars and restaurants, clothing, residential construction, and even a service such as telecommunications. The limitations in real European data made heroic assumptions necessary to operationalize the model. One conclusion, therefore, is that provision of data in Europe urgently needs to be improved, at least to the level of sector detail currently available for the United States and Japan.     

Identification of bacteria with beta-galactosidase activity in faeces from lactase non-persistent subjects

Previous studies suggest that, besides the maldigestion of lactose in the small intestine, the colonic processing of lactose might play a role in lactose intolerance. beta-Galactosidase is the bacterial enzyme which catalyzes the first step of lactose fermentation in the colon. We propose a practical method to differentiate and identify bacteria with beta-galactosidase activity in faeces which combines a colony-lift filter assay with X-gal (5-bromo-4-chloro-3-indolyl-beta-d-galactopyranoside) as substrate for differentiation and the fluorescent in situ hybridization technique for identification. The method was applied to faeces from lactase non-persistent subjects. After 28 subjects had undergone one glucose and two lactose challenges, consistent intolerant (n=5) and tolerant (n=7) groups were defined according to their symptom scores. Of the 28 faecal samples, 80.6% (mean, SD: 12.1, range: 47.8-100%) of the total cultured bacteria were found to possess beta-galactosidase activity, which indicates that the bacterial beta-galactosidase is abundant in the colon. The tolerant and intolerant groups did not differ in the percentage or composition of the bacteria with beta-galactosidase activity or beta-galactosidase activity in faeces. Results suggest that the percentage or composition of the bacteria with beta-galactosidase activity in faeces do not play a role in lactose intolerance.